I propose to isolate and characterize the deatiled structure of precursors of alfatoxins, one of the most potent mutagen/carcinogin molecules known. Kinetic pulse-labeling techniques were successfully adapted for use with Aspergillus versicolor, the validity of this approach for isolating precursor molecules being confirmed by tracing known intermediates. I have extended this approach to A. parasiticus, the toxigenic strain which produces aflatoxins. New techniques of radial thin-layer chromatography have also been developed and cross-feeding of unknown intermediates between various mutant combinations has worked exceedingly well. Early pathway mutants were isolated using a genetic background which allows the convenient selection of averufin-negative mutants. The latter mutants were derived as secondary mutants from a parent stock of A. parasiticus which accumulates averufin (i.e., has a post-averufin, block in aflatoxin biosynthesis). A new early precursor, denoted pre-averufin, has been isolated. Complete structural determination of pre-averufin will be carried out through the application of spectrascopic techniques (1H-NMR, 13C-NMR and mass spectrometry). Isolation and proof-of-structure data will also be sought for other precursors using both early-and lateblocked mutants already obtained in our laboratory following mutagenesis of A. parasiticus.